This grant proposal focuses on the critical role of the transcription factor, human achaete-scute homolog-1 (hASH1) in the evolution of small cell lung cancer (SCLC). Unlike the other three major types of lung cancer, SCLC is distinguished by expression of neural and neuroendocrine (NE) features. HASH1 is a human counterpart to the Drosophila achaete-scute complex, a conserved family of basic helix loop helix transcription factors that are essential for commitment of primitive neuroblasts from undifferentiated precursors. In mammalian development, ASH1 (termed MASH1 in rodents) is essential for normal differentiation of sympathoadrenal, enteric, olfactory, retinal, and brain sympathetic neurons. We have recently shown that ASH1 in the normal fetal lung is restricted to lung NE cells. Furthermore, transgenic knockout of ASH1 results in a complete failure of lung NE cells to differentiate and causes fatal respiratory insufficiency at birth. Preliminary studies from lung injury models suggest an important role for hASH1 in the recruitment of hyperplastic lung NE cells. Across a broad spectrum of lung cancer phenotypes, hASH1 expression is strictly concordant with NE phenotypic markers. We found that depletion of hASH1 protein from cultured SCLC cells resulted in a comparable reduction of NE marker expression, and an increase in apoptotic fraction. We have studied a new transgenic model in which hASH1 is targeted to lung Clara cells, using a cell-specific CC10 promoter. These mice develop progressive hyperplasia of their distal airway involving the CC10-reactive cells. Remarkably, hASH1 cooperates potently with the SV40 T-Antigen to promote malignant transformation and NE tumors in these target cells. Collectively, these findings indicate that hASH1 may be critical in the evolution of SCLC from lung NE precursor cells. Our proposed series of studies now seek to determine: 1) How hASH1 and related proneural transcription factors function in novel models of lung carcinogenesis; 2) What are the transcriptional targets of hASH1, relevant to the evolution of lung cancer, and 3) How hASH1 regulation, via the Notch pathway, may be altered in SCLC cells. These studies will provide a more comprehensive understanding of hASH1 action in SCLC pathogenesis, potentially leading to exploitation of specific vulnerabilities of SCLC tumors in novel treatment strategies.